One of the important unresolved issues in physiological shock is understanding the mechanism leading to formation of an inflammatory cascade. Shock is accompanied by cell activation in the microcirculation, leukocyte infiltration, cell dysfunction, apoptosis and organ failure. We recently obtained evidence that pancreatic enzymes serve as a powerful source for generation of humoral inflammatory mediators in the ischemic intestine. Blockade of pancreatic enzymes in the lumen of the ischemic intestine leads to high levels of protection against inflammation and multi-organ failure. Our results point to an important role for pancreatic serine proteinases. In accordance with this evidence we hypothesize that pancreatic digestive enzymes in the intestine can escape across the brush border cell barrier during ischemia and thereafter produce humoral inflammatory mediators by digestion of extracellular matrix proteins and other cellular components. Our Specific Aims are (1) to identify specific pancreatic enzyme activities which contribute to humoral microvascular activator production in hemorrhagic shock by enzyme blockade in the lumen of the ischemic intestine and by introduction of purified pancreatic enzymes into the lumen of the small intestine; (2) to examine the pancreatic enzyme localization in the tissue before and after intestinal ischemia and determine the production of inflammatory mediator production in the interstitial space of the small intestine; (3) to examine molecular mechanisms for initiation of inflammation in a peripheral organ after activator production by pancreatic enzymes in the intestine; and (4) to purify and characterize selected inflammatory mediators produced by pancreatic enzymes from homogenates of pancreas as an ubiquitous source of inflammatory mediators and from digests of purified extracellular matrix proteins (including fibronectin, vitronectin, and collagen I, III and IV) with serine proteinases. We will study, using state of the art digital in-vivo microscopy, transport and action of pancreatic enzymes in combination with biochemical identification of inflammatory mediators in the rat shock model. Biochemical identification of the inflammatory mediators is facilitated by availability of large quantities of starting material from harvested rat and pig pancreatic tissue. Understanding the trigger mechanisms for inflammation in shock will lead to new treatment modalities in man.